Elimination of HIV-1 plasmid DNA from virus samples obtained from transfection by calcium–phosphate co-precipitation

Molecular genetics is a powerful tool to analyze the replication cycle of human immunodeficiency virus type 1 (HIV-1). Culture fluids obtained from HIV-1 plasmid-transfected cells by calcium–phosphate co-precipitation were treated with ethyleneglycol bis (β-aminoethylether)- N, N, N′, N′-tetraacetic...

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Bibliographic Details
Published in:Journal of virological methods 2000-10, Vol.90 (1), p.99-102
Main Authors: Koh, Kyu-Bom, Fujita, Mikako, Adachi, Akio
Format: Article
Language:English
Subjects:
Biological and medical sciences
Calcium - chemistry
calcium phosphate
Calcium–phosphate coprecipitation
Cell Line
Chemical Precipitation
Deoxyribonucleases - metabolism
DNA, Viral - chemistry
DNA, Viral - physiology
EGTA
Egtazic Acid - pharmacology
Fundamental and applied biological sciences. Psychology
HIV-1
HIV-1 - genetics
HIV-1 - isolation & purification
HIV-1 - physiology
Human immunodeficiency virus 1
Humans
Microbiology
Phosphates - chemistry
Plasmids - genetics
Polymerase Chain Reaction
Techniques used in virology
Transfection
Virology
Virus Replication
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Summary:Molecular genetics is a powerful tool to analyze the replication cycle of human immunodeficiency virus type 1 (HIV-1). Culture fluids obtained from HIV-1 plasmid-transfected cells by calcium–phosphate co-precipitation were treated with ethyleneglycol bis (β-aminoethylether)- N, N, N′, N′-tetraacetic acid (EGTA) and DNase I to obtain HIV-1 stocks virtually free of input plasmid DNAs. Even after amplification by polymerase chain reaction (PCR), no plasmid DNA was detected in cells following infection with EGTA/DNase I-treated virus samples. This method is particularly useful for the examination of the early replication phase of HIV-1 by PCR.
ISSN:0166-0934
1879-0984
DOI:10.1016/S0166-0934(00)00224-X